SBB10Ntbk-Mesa
2/17/2010
Received Oligos: KRM001, KRM002, KRM003, KRM004
Diluted all oligos to a 100μM concentration
Made 10μM dilutions of KRM001 and KRM 002
9μl H2O 1μl 100μM oligo
Prepared one wobble and one PCR reaction, KRM16 and KRM27 respectively.
Wobble Reaction (KRM16) 29μl ddH2O 5μl 2 5μl dNTPs 5μl KRM003 (100μM) 5μl KRM004 (100μM) 0.75μl 1 PCR Reaction (KRM27) 24μl ddH2O 3.3μl 2 3.3μl dNTPs 1μl KRM003 (10μM) 1μl KRM004 (10μM) 0.5μl 1 0.5μl Template DNA (pEC22-CA42)
2/22/2010
Zymo Clean Up on PCR (KRM27)
Elute in 33μl
Small Fragment Zymo on Wobble (KRM16)
Elute in 50μl
Ran an Analytical Gel for Purified PCR (KRM27) and Wobble (KRM16) samples
3μl Purified PCR (KRM27) 7μl Loading Buffer 3μl Purified Wobble (KRM16) 7μl Loading Buffer
Purified Samples were stored in140L Box A
Gel Results
KRM27 expected size is 987bp (Second to far right lane)
KRM16 expected size is 115bp (Far right lane)
2/24/2010
PCR EcoRI/BamHI Digest on Purified PCR (KRM27)
Remaining Purified PCR product in tube labeled->KRM27, Purified PCR product, 2/22/10
Wobble EcoRI/BamHI Digest on Purified Wobble (KRM16)
All of Purified Wobble product was used in digest
Small Fragment Zymo on Wobble Digest (KRM16 Digest)
Elute in 50μl Wobble digest product in tube labeled->KRM16 Digest, Purified Digest, 2/24/10
Ran a Preparative and an Analytical Gel for PCR Digest (KRM27 Digest) and Wobble Digest (KRM16 Digest) samples, respectively
9μl PCR Digest (KRM27 Digest) 1μl 6X Loading Buffer 3μl Wobble Digest (KRM16 Digest) 7μl Loading Buffer
Cut out PCR Digest from preparative gel, place in 1.5ml tube Add 600μl ADB buffer @ 55°C
Gel Results
KRM16 Digest expected size is 115bp (Far right lane)
Samples were stored in140L Box A
